Rat NSC have been tested for proliferation and differentiation potential up to passage 3 after thawing and we have seen around 300 million cells from 1 vial after the 3rd passage. We have not expanded the cells beyond the third passage, but we encourage you to try expanding further.

Cell purity was measured by the expression of NSC marker of Nestin and devoid of expression of differentiated marker. More than 75% of the cells were positive to Nestin and expressions of the differentiated markers were less than 10%.

The doubling time for the cells is around 20-30 hours. This tends to increase with passage number.

9. Cells do not attach to the CELLStartTM coated plate but float to form a sphere. Is this normal?

Please ensure that the buffer used for CELLStart has Calcium and Magnesium. Please review the product insert for our recommended buffer. If the right buffer is used, increase the concentration of CELLStart up to 1:50 dilution rate. If CellStart with our recommended buffer does not work, some users had success with Poly-L-Orinithin coated plates with StemPro NSC-SFM. If you use Poly-L-Orinithin, please perform overnight incubation.

NSC is a valuable source not only for neuroscience but also for clinical use to treat neurodegenerative disease or neurological disorder. These cells can be used for a wide range of differentiation studies.

13. Where can I find more detailed protocol for directed differentiation?

Please contact our technical support team at Techsupport@invitrogen.com for the details. Our product manual will however guide you in the protocol for spontaneous differentiation. All three lineages can be obtained from spontaneous differentiation. However the % of the various cell types (neurons, astrocytes and oligodendrocytes) will be few and will depend on the passage number.

Any format of culture vessel (e.g. flask, 6-well plates, etc) can be used. Seed cells at a density of 5 x 104 cells/cm2. Depending on the surface area of your culture vessel, calculate how many cells you will need to have on hand. Example: For one 60 mm dish, you will need 5 x 104 cells/cm2 x 20 cm2 = 1 x 106 cells total.

GIBCO Rat Fetal NSC’s are derived from a mixed pool of embryos; therefore, cells will be derived from rats of both sexes.

16. Is there any difference between NSCs isolated from cortex vs hippocampus?

Cortical NSC’s contain a greater mixture of cells and have different neurotransmitters. They have a wider set of applications. Fewer NSCs can be obtained from hippocampal tissue, but the cells are more pure. They are used by a smaller hippocampal focused audience.

17. What are the estimated % neurons, astrocytes, and oligodendrocytes that we might get from using a spontaneous differentiation protocol vs. a directed differentiation protocol?